鄭安理2006-07-262018-07-112006-07-262018-07-112005http://ntur.lib.ntu.edu.tw//handle/246246/23677皮質類固醇除本身對於某些血液腫瘤具細胞毒性之外，也常與抗癌化學藥物併用以治 療因化學藥物引起之噁心、嘔吐及過敏反應等副作用。雖然類固醇己被証實可以影響多種 細胞之重要訊息傳遞徑路，其中有些與癌細胞抗藥性有關。然而我們對類固醇類藥物對於 與一般癌細胞生長以及化學藥物感受性可能產生之影響仍所知極少。釐清這個問題對臨床 腫瘤治療將會有重要影響。 我們隨機選擇了十四株癌細胞株有系統地進行研究以解答這個問題。Dexamethasone （DEX）被選為皮質類固醇代表藥物。我們發現： 1.DEX 確實對癌細胞株（十四株之中的七株）的生長以及化學藥物感受性有影響。DEX 對 癌細胞的影響呈現異質性而且似乎是彼此互斥的。DEX（0.01~1.0uM）抑制四株細胞的 生長(MCF-7 ，MCF/MXR1 ，MCF/TPT300 及HeLa 細胞)，提高了一株細胞對cisplatin 的化 學藥物感受性(SiHa)，並降低兩株細胞對cisplatin,doxorubicin,5FU,及taxol 的 化學藥物感受性(H460 及Hep3B)。 2.此影響是皮質類固醇受體—依賴性的。因為DEX 只有在含有高濃度皮質類固醇受體(≧ 2.1x10 4 /細胞)的七株細胞才有影響。在其他七株不受DEX 影響的細胞中，有五株細胞 皮質類固醇受體濃度範圍僅在2.0~5.7x10 3 /細胞之間。而另二株含有高濃度皮質類固醇 受體但不受DEX 影響的細胞中(TW01,TW04)，我們發現其皮質類固醇受體不具有功能。 3.DEX 在SiHa 細胞所造成的化學藥物致敏感效應與其對NF-kB 的調控有著高度相關。透 過轉殖含有dominant negative IkB 的plasmid 進入SIHa cells 中以抑制NFkB 活性， 我們發現原先DEX 提高SiHa 細胞對cisplatin 的化學藥物感受性的現象消失。而在我 們進一步的研究發現在SiHa 細胞中皮質類固醇受體確實會與NFkB 有直接作用(dirrect protein-protein interaction)。 4.我們另外發現DEX 會直接作用於MCF7 細胞的p21 promoter 上造成p21 的向上調控， 表現量增加進而造成細胞週期G1 arrest,抑制癌細胞的生長。 5.為了尋找可能造成DEX 對癌細胞株的生長以及化學藥物感受性異質性影響的機轉或決 定性因子(determinant factor)，我們研究皮質類固醇受體共同協調因子(steroid receptor co-regulator)在這些細胞的表現。我們發現這些對DEX 有著不同反應的癌細 胞中皮質類固醇受體共同協調因子(steroid receptor co-regulator)的表現各有不 同。而且其中有許多共同協調因子之表現量本身亦受到皮質類固醇受體所影響。 6.利用即時定量RT-PCR 測定，以及利用免疫組織化學染色檢驗一百多位病患的(肺癌、乳 癌、子宮頸癌)癌細胞檢體我們發現在不同病人的癌細胞中的皮質類固醇受體含量有高 有低。我們推測臨床上有一定比例之癌細胞含有高濃度皮質類固醇受體，並可能對DEX 有感受性。 利用免疫組織染色測定癌病患癌細胞檢體，我們發現： 1. 在不同病人的癌細胞中的皮質類固醇受體含量有高有低。我們推測臨床上有一定比例 之癌細胞含有高濃度皮質類固醇受體，並可能對DEX 有感受性。 2. 在非小細胞肺癌的患者中，癌細胞中的皮質類固醇受體含量較高者其臨床預後較佳Objectives: Glucocorticoids (GCs) are commonly co-administered with anti-cancer drugs such as cisplatin to prevent drug-induced allergic reaction, nausea, and vomiting. But little is known regarding the effects of GCs on the growth and chemosensitivity of common carcinomas cells. Methods: Fourteen carcinoma cell lines representing breast (MCF-7, MCF-7/MXR1, MCF-7/TPT300), gastric (AGS, N87,SNU1), lung (H460), cervical (SiHa, HeLa, Caski), liver (Hep3B, Hut7), and nasopharyngeal (NPC-TW01, NPC-TW04) cancer were selected to assess the effects of dexamethasone (DEX) on the cell growth and cisplatin chemosensitivity of common human cancers. Immunohistochemical stain of tissues and cells were done by PA1-511A, an anti-GR monoclonal antibody. Results: DEX had mutually exclusive effects on either growth or cisplatin sensitivity in 7 of the 14 cell lines. DEX inhibited cell growth of 4 (MCF-7, MCF-7/MXR1, MCF-7/TPT300, and HeLa), increased cisplatin cytotoxicity of one (SiHa), and decreased cisplatin cytotoxicity of 2 (H460 and Hep3B) cells lines. Although the effect of DEX on these carcinoma cells was unexpectedly diverse, it remained GC receptor (GCR) dependent. The GCR contents of the 7 cell lines affected by DEX were significantly higher than those of the other 7 cell lines unaffected by DEX (5.2±2.5×10 4 vs 1.3±1.4×10 4 , P=0.005).Only two DEX-unresponsive cell lines (NPC-TW01 and NPC-TW04) had GCR contents at the high range as those of the 7 DEX-responsive cell lines. On further examination, the function of the endogenous GCR of these two cell lines was found to be impaired. Further, transfection and expression of a vector encoding GCR to AGS, a GCR low-expressing and GC non-responsive cell line, increased its susceptibility to DEX manifested as an increased resistance toward cisplatin. The cytotoxicity-enhancing effect of GC in SiHa cells correlated well with its effect on abrogating the cisplatin-induced activation of NF-κ B. Expression of a dominant-negative truncated I κ B α gene in SiHa cells completely abolished the cytotoxicity-enhancing effect of DEX. Conclusions: GCs may affect growth or chemosensitivity of carcinoma cells containing high concentration of functional GCR. Although the effects are heterogeneous and currently unpredictable , our data underscore the importance of clarifying the impact on tumor control by the co-administed GCs to carcinoma patients receiving chemotherapy. The expressions of the steroid receptor co-regulators of these cells are examined. Correlation with the difference between expression of the coregulators and DEX responsiveness were also examined. Furthermore, the expressions of some of the co-regulators were influenced by the GCR. In human cancer samples, we demonstrated that some of the breast, lung and uterine cervical cancer do express high level of GCR. In MCF7 cells, we found DEX induced p21 up-regulation and caused G1 phase arrest of MCF-7. Addition of excess amounts of a structure homologue of DEX, RU486, completely abolished the growth suppression effect of DEX, suggesting that DEX act via GCR-related signal transduction pathways. Furthers, DEX has no effect on the growth of MCF-7/GCR(-), an MCF-7 subclone contains vary low levels of GCR (<1x10 3 /cell). Compared with MCF-7, MCF-7/GCR(-) contains no detectable level of CBP300, HDAC1, and significantly lower levels of NCOR1, TIF2, GCN5L2, and ARA70. Transfection of GCR RNAi to MCF-7 cells also resulted in no detectable level of CBP300, HDAC1, and significantly lowers levels of NCOR1, TIF2, GCN5L2, and ARA70. Transfection of human GCR to MCF-7/GCR(-) restored the expression of GCR and all these co-regulators and sensitivity to DEX in MCF-7/GCR(-) cells. Chromosome IP with anti-GCR antibody and PCR study showed positive result with TIF-2, imply the possibility of direct regulation of TIF-2 expression by GCR. Immunohistochemical studies of human cancer tissues showed that 5 of the 45 (11.1%) breast cancer and 43 of the 85 (50.6%) non-small cell lung cancer had high GR contents at the ranges of the glucocorticoid-responsive carcinoma cell lines. High GR expression was detected in 51% of the tumor specimens. The difference in tumor GR expression was not associated with cell type, gender, age, or stage. The outcome was significantly superior for patients whose tumor showed high GR expression compared to those with either low expression or non-expression. The median progression-free survival was 8.0 vs. 5.6 months (p=0.039) and overall survival was 18.1 vs. 10.2 months, (p=0.003), respectively. Almost all these patients have received GC as antiemetics or allergic preventive treatment during chemotherapy courses, therefore, the effect of GC on the chemosensitivity in vivo was not evaluable. Conclusions: GCs may affect growth or chemosensitivity of carcinoma cells containing high concentration of functional GCR. Although the effects are heterogeneous and currently unpredictable, our data underscore the importance of clarifying the impact on tumor control by the co-administed GCs to carcinoma patients receiving chemotherapy. It is mandatory to identify the molecular and cellular markers that help predict the diverse effect of GCs on carcinoma cells.application/pdf492053 bytesapplication/pdfzh-TW國立臺灣大學醫學院內科皮質類固醇癌細胞化學藥物感受性GlucocorticoidsGlucocorticoid receptorCarcinomaCell growthChemosensitivityDrug resistance[SDGs]SDG3reporthttp://ntur.lib.ntu.edu.tw/bitstream/246246/23677/1/932314B002006.pdf